Pyrazolopyrimidines as cyclin dependent kinase inhibitors

ABSTRACT

In its many embodiments, the present invention provides a novel class of pyrazolo[1,5-a]pyrimidine compounds as inhibitors of cyclin dependent kinases, methods of preparing such compounds, pharmaceutical compositions containing one or more such compounds, methods of preparing pharmaceutical formulations comprising one or more such compounds, and methods of treatment, prevention, inhibition, or amelioration of one or more diseases associated with the CDKs using such compounds or pharmaceutical compositions.

This application claims benefit of priority from U.S. provisional patentapplication Ser. No. 60/408,029 filed Sep. 4, 2002.

FIELD OF THE INVENTION

The present invention relates to pyrazolo[1,5-a]pyrimidine compoundsuseful as protein kinase inhibitors, pharmaceutical compositionscontaining the compounds, and methods of treatment using the compoundsand compositions to treat diseases such as, for example, cancer,inflammation, arthritis, viral diseases, neurodegenerative diseases suchas Alzheimer's disease, cardiovascular diseases, and fungal diseases.

BACKGROUND OF THE INVENTION

The cyclin-dependent kinases (CDKs) are serine/threonine proteinkinases, which are the driving force behind the cell cycle and cellproliferation. Individual CDK's, such as, CDK1, CDK2, CDK3, CDK4, CDK5,CDK6 and CDK7, CDK8 and the like, perform distinct roles in cell cycleprogression and can be classified as either G1, S, or G2M phase enzymes.Uncontrolled proliferation is a hallmark of cancer cells, andmisregulation of CDK function occurs with high frequency in manyimportant solid tumors. CDK2 and CDK4 are of particular interest becausetheir activities are frequently misregulated in a wide variety of humancancers. CDK2 activity is required for progression through G1 to the Sphase of the cell cycle, and CDK2 is one of the key components of the G1checkpoint. Checkpoints serve to maintain the proper sequence of cellcycle events and allow the cell to respond to insults or toproliferative signals, while the loss of proper checkpoint control incancer cells contributes to tumorgenesis. The CDK2 pathway influencestumorgenesis at the level of tumor suppressor function (e.g. p52, RB,and p27) and oncogene activation (cyclin E). Many reports havedemonstrated that both the coactivator, cyclin E, and the inhibitor,p27, of CDK2 are either over—or underexpressed, respectively, in breast,colon, nonsmall cell lung, gastric, prostate, bladder, non-Hodgkin'slymphoma, ovarian, and other cancers. Their altered expression has beenshown to correlate with increased CDK2 activity levels and poor overallsurvival. This observation makes CDK2 and its regulatory pathwayscompelling targets for the development years, a number of adenosine5′-triphosphate (ATP) competitive small organic molecules as well aspeptides have been reported in the literature as CDK inhibitors for thepotential treatment of cancers. U.S. Pat. No. 6,413,974, col. 1, line23- col. 15, line 10 offers a good description of the various CDKs andtheir relationship to various types of cancer.

CDK inhibitors are known. For example, flavopiridol (Formula I) is anonselective CDK inhibitor that is currently undergoing human clinicaltrials, A. M. Sanderowicz et al, J. Clin. Oncol. (1998) 16, 2986–2999.

Other known inhibitors of the CDKs include, for example, olomoucine (J.Vesely et al, Eur. J. Biochem., (1994) 224, 771–786) and roscovitine (I.Meijer et al, Eur. J. Biochem., (1997) 243, 527–536). U.S. Pat. No.6,107,305 describes certain pyrazolo[3,4-b] pyridine compounds as CDKinhibitors. An illustrative compound from the '305 patent has theFormula II:

K. S. Kim et al, J. Med. Chem. 45 (2002) 3905–3927 and WO 02/10162disclose certain aminothiazole compounds as CDK inhibitors.

Pyrazolopyrimidines are known. For Example, WO92/18504, WO02/50079,WO95/35298, WO02/40485, EP94304104.6, EP0628559 (equivalent to U.S. Pat.Nos. 5,602,136, 5,602,137 and 5,571,813), U.S. Pat. No. 6,383,790, Chem.Pharm. Bull., (1999) 47 928, J. Med. Chem., (1977) 20, 296, J. Med.Chem., (1976) 19 517 and Chem. Pharm. Bull., (1962) 10 620 disclosevarious pyrazolopyrimidines.

There is a need for new compounds, formulations, treatments andtherapies to treat diseases and disorders associated with CDKs. It is,therefore, an object of this invention to provide compounds useful inthe treatment or prevention or amelioration of such diseases anddisorders.

SUMMARY OF THE INVENTION

In its many embodiments, the present invention provides a novel class ofpyrazolo[1,5-a]pyrimidine compounds as inhibitors of cyclin dependentkinases, methods of preparing such compounds, pharmaceuticalcompositions comprising one or more such compounds, methods of preparingpharmaceutical formulations comprising one or more such compounds, andmethods of treatment, prevention, inhibition or amelioration of one ormore diseases associated with the CDKs using such compounds orpharmaceutical compositions.

In one aspect, the present application discloses a compound, orpharmaceutically acceptable salts or solvates of said compound, saidcompound having the general structure shown in Formula III:

wherein:

R is an unsubstituted aryl or aryl substituted with one or more moietieswhich moieties can be the same or different, each moiety beingindependently selected from the group consisting of halogen, CN, —OR⁵,SR⁵, —CH₂OR⁵, —C(O)R⁵, —SO₃H, —S(O₂)R⁶, —S(O₂)NR⁵R⁶, —NR⁵R⁶, —C(O)NR⁵R⁶,—CF₃, —OCF₃ and heterocyclyl;

R² is selected from the group consisting of R⁹, alkyl, alkynyl,alkynylalkyl, cycloalkyl, —CF₃, —C(O₂)R⁶, aryl, arylalkyl,heteroarylalkyl, heterocyclyl, alkyl substituted with 1–6 R⁹ groupswhich groups can be the same or different with each R⁹ beingindependently selected, aryl substituted with 1–3 aryl or heteroarylgroups which can be the same or different and are independently selectedfrom phenyl, pyridyl, thiophenyl, furanyl and thiazolo groups,

R³ is selected from the group consisting of H, halogen, —NR⁵R⁶,—C(O)NR⁵R⁶, alkyl, alkynyl, cycloalkyl, aryl, arylalkyl, heterocyclyl,heterocyclylalkyl, heteroaryl and heteroarylalkyl,

wherein each of said alkyl, cycloalkyl, aryl, arylalkyl, heterocyclyl,heterocyclylalkyl, heteroaryl and heteroarylalkyl for R³ and theheterocyclyl moieties whose structures are shown immediately above forR³ can be substituted or optionally independently substituted with oneor more moieties which can be the same or different, each moiety beingindependently selected from the group consisting of halogen, alkyl,aryl, cycloalkyl, CF₃, CN, —OCF₃, —(CR⁴R⁵)_(n)OR⁵, —OR⁵, —NR⁵R⁶,—(CR⁴R⁵)_(n)NR⁵R⁶, —C(O₂)R⁵, —C(O)R⁵, —C(O)NR⁵R⁶, —SR⁶, —S(O₂)R⁶,—S(O₂)NR⁵R⁶, —N(R⁵)S(O₂)R⁷, —N(R⁵)C(O)R⁷ and —N(R⁵)C(O)NR⁵R⁶;

R⁴ is H, halo or alkyl;

R⁵ is H or alkyl;

R⁶ is selected from the group consisting of H, alkyl, aryl, arylalkyl,cycloalkyl, heterocyclyl, heterocyclylalkyl, heteroaryl, andheteroarylalkyl, wherein each of said alkyl, aryl, arylalkyl,cycloalkyl, heterocyclyl, heterocyclylalkyl, heteroaryl, andheteroarylalkyl can be unsubstituted or optionally substituted with oneor more moieties which can be the same or different, each moiety beingindependently selected from the group consisting of halogen, alkyl,aryl, cycloalkyl, heterocyclylalkyl, CF₃, OCF₃, CN, —OR⁵, —NR⁵R¹⁰,—N(R⁵)Boc, —(CR⁴R⁵)_(n)OR⁵, —C(O₂)R⁵, —C(O)R⁵, —C(O)NR⁵R¹⁰, —SO₃H,—SR¹⁰, —S(O₂)R⁷, —S(O₂)NR⁵R¹⁰, —N(R⁵)S(O₂)R⁷, —N(R⁵)C(O)R⁷ and—N(R⁵)C(O)NR⁵R¹⁰;

R¹⁰ is selected from the group consisting of H, alkyl, aryl, arylalkyl,cycloalkyl, heterocyclyl, heterocyclylalkyl, heteroaryl, andheteroarylalkyl, wherein each of said alkyl, aryl, arylalkyl,cycloalkyl, heterocyclyl, heterocyclylalkyl, heteroaryl, andheteroarylalkyl can be unsubstituted or optionally substituted with oneor more moieties which can be the same or different, each moiety beingindependently selected from the group consisting of halogen, alkyl,aryl, cycloalkyl, heterocyclylalkyl, CF₃, OCF₃, CN, —OR⁵, —NR⁴R⁵,—N(R⁵)Boc, —(CR⁴R⁵)_(n)OR⁵, —C(O₂)R⁵, —C(O)N R⁴R⁵, —C(O)R⁵, —SO₃H, —SR⁵,—S(O₂)R⁷, —S(O₂)NR⁴R⁵, —N(R⁵)S(O₂)R⁷, —N(R⁵)C(O)R⁷ and —N(R⁵)C(O)NR⁴R⁵;

-   -   or optionally (i) R⁵ and R¹⁰ in the moiety —NR⁵R¹⁰, or (ii) R⁵        and R⁶ in the moiety —NR⁵R⁶, may be joined together to form a        cycloalkyl or heterocyclyl moiety, with each of said cycloalkyl        or heterocyclyl moiety being unsubstituted or optionally        independently being substituted with one or more R⁹ groups;

R⁷ is selected from the group consisting of alkyl, cycloalkyl, aryl,heteroaryl, arylalkyl and heteroarylalkyl, wherein each of said alkyl,cycloalkyl, heteroarylalkyl, aryl, heteroaryl and arylalkyl can beunsubstituted or optionally independently substituted with one or moremoieties which can be the same or different, each moiety beingindependently selected from the group consisting of halogen, alkyl,aryl, cycloalkyl, CF₃, OCF₃, CN, —OR⁵, —NR⁵R¹⁰, —CH₂OR⁵, —C(O₂)R⁵,—C(O)NR⁵R¹⁰, —C(O)R⁵, —SR¹⁰, —S(O₂)R¹⁰, —S(O₂)NR⁵R¹⁰, —N(R⁵)S(O₂)R¹⁰,—N(R⁵)C(O)R¹⁰ and —N(R⁵)C(O)NR⁵R¹⁰;

R⁸ is selected from the group consisting of R⁶, —C(O)NR⁵R¹⁰,—S(O₂)NR⁵R¹⁰, —C(O)R⁷ and —S(O₂)R⁷;

R⁹ is selected from the group consisting of halogen, CN, —NR⁵R¹⁰,—C(O₂)R⁶, —C(O)NR⁵R¹⁰, —OR⁶, —SR⁶, —S(O₂)R⁷, —S(O₂)NR⁵R¹⁰,—N(R⁵)S(O₂)R⁷, —N(R⁵)C(O)R⁷and —N(R⁵)C(O)NR⁵R¹⁰;

m is 0 to 4, and

n is 1 to 4,

with the following provisos: (i) that when R is an unsubstituted phenyl,then R² is not alkyl, —C(O₂)R⁶, aryl or cycloalkyl, and (ii) that when Ris a phenyl substituted with a hydroxyl group, then R² is halogen only.

The compounds of Formula III can be useful as protein kinase inhibitorsand can be useful in the treatment and prevention of proliferativediseases, for example, cancer, inflammation and arthritis. They may alsobe useful in the treatment of neurodegenerative diseases suchAlzheimer's disease, cardiovascular diseases, viral diseases and fungaldiseases.

DETAILED DESCRIPTION

In an embodiment, the present invention disclosespyrazolo[1,5-a]pyrimidine compounds which are represented by structuralFormula III, or a pharmaceutically acceptable salt or solvate thereof,wherein the various moieties are as described above.

In another embodiment, R is an unsubstituted aryl or aryl substitutedwith one or moieties which moieties can be the same or different witheach moiety being independently selected from the group consisting ofhalogen, CN, —OR⁵, —S(O₂)NR⁵R⁶, —SO₃H, CH₂OR⁵, —S(O₂)R⁶, —C(O)NR⁵R⁶,—CF₃, —OCF₃ and heterocyclyl.

In another embodiment, R² is halogen, CF₃, CN, lower alkyl andcycloalkyl.

In another embodiment, R³ is H, unsubstituted aryl, unsubstitutedheteroaryl, aryl substituted with one or more moieties selected from thegroup consisting of halogen, CN, —OR⁵, CF₃, —OCF₃, lower alkyl andcycloalkyl, heteroaryl substituted with one or more moieties selectedfrom the group consisting of halogen, CN, —OR⁵, CF₃, —OCF₃, alkyl andcycloalkyl, and heterocyclyl.

In another embodiment, R⁴ is H or lower alkyl.

In another embodiment, R⁵ is H or lower alkyl.

In another embodiment, n is 1 or 2.

In an additional embodiment, R is unsubstituted phenyl.

In an additional embodiment, R is phenyl substituted with one or moremoieties selected from the group consisting of F, Cl, Br, CN, —SO₃H,—S(O₂)NR⁵R⁶, —S(O₂)CH₃, —OH, CF₃ and morpholinyl.

In an additional embodiment, R² is F, Cl, Br, CF₃, lower alkyl,cyclopropyl, cyclobutyl or cyclopentyl.

In an additional embodiment, R³ is H, aryl wherein said aryl can beunsubstituted or optionally substituted with one or more moieties whichcan be the same or different, each moiety being independently selectedfrom the group consisting of F, Cl, Br, CF₃, lower alkyl, methoxy andCN, alkyl, heteroaryl, heterocyclyl or heterocyclyl substituted with atleast one hydroxyalkyl.

In an additional embodiment, R³ is 2-fluorophenyl, 2-chlorophenyl,2,3-dichlorophenyl, 2-methylphenyl, 2methoxyphenyl,

In an additional embodiment, R³ is:

In an additional embodiment, R⁴ is H.

In an additional embodiment, R⁵ is H.

An inventive group of compounds are shown in Table 1.

TABLE 1

As used above, and throughout this disclosure, the following terms,unless otherwise indicated, shall be understood to have the followingmeanings:

“Patient” includes both human and animals.

“Mammal” means humans and other mammalian animals.

“Alkyl” means an aliphatic hydrocarbon group which may be straight orbranched and comprising about 1 to about 20 carbon atoms in the chain.Preferred alkyl groups contain about 1 to about 12 carbon atoms in thechain. More preferred alkyl groups contain about 1 to about 6 carbonatoms in the chain. Branched means that one or more lower alkyl groupssuch as methyl, ethyl or propyl, are attached to a linear alkyl chain.“Lower alkyl” means a group having about 1 to about 6 carbon atoms inthe chain which may be straight or branched. The term “substitutedalkyl” means that the alkyl group may be substituted by one or moresubstituents which may be the same or different, each substituent beingindependently selected from the group consisting of halo, alkyl, aryl,cycloalkyl, cyano, hydroxy, alkoxy, alkylthio, amino, —NH(alkyl),—NH(cycloalkyl), —N(alkyl)₂, carboxy and —C(O)O-alkyl. Non-limitingexamples of suitable alkyl groups include methyl, ethyl, n-propyl,isopropyl and t-butyl.

“Alkynyl” means an aliphatic hydrocarbon group containing at least onecarbon-carbon triple bond and which may be straight or branched andcomprising about 2 to about 15 carbon atoms in the chain. Preferredalkynyl groups have about 2 to about 12 carbon atoms in the chain; andmore preferably about 2 to about 4 carbon atoms in the chain. Branchedmeans that one or more lower alkyl groups such as methyl, ethyl orpropyl, are attached to a linear alkynyl chain. “Lower alkynyl” meansabout 2 to about 6 carbon atoms in the chain which may be straight orbranched. Non-limiting examples of suitable alkynyl groups includeethynyl, propynyl, 2-butynyl and 3-methylbutynyl. The term “substitutedalkynyl” means that the alkynyl group may be substituted by one or moresubstituents which may be the same or different, each substituent beingindependently selected from the group consisting of alkyl, aryl andcycloalkyl.

“Aryl” means an aromatic monocyclic or multicyclic ring systemcomprising about 6 to about 14 carbon atoms, preferably about 6 to about10 carbon atoms. The aryl group can be optionally substituted with oneor more “ring system substituents” which may be the same or different,and are as defined herein. Non-limiting examples of suitable aryl groupsinclude phenyl and naphthyl.

“Heteroaryl” means an aromatic monocyclic or multicyclic ring systemcomprising about 5 to about 14 ring atoms, preferably about 5 to about10 ring atoms, in which one or more of the ring atoms is an elementother than carbon, for example nitrogen, oxygen or sulfur, alone or incombination. Preferred heteroaryls contain about 5 to about 6 ringatoms. The “heteroaryl” can be optionally substituted by one or more“ring system substituents” which may be the same or different, and areas defined herein. The prefix aza, oxa or thia before the heteroarylroot name means that at least a nitrogen, oxygen or sulfur atomrespectively, is present as a ring atom. A nitrogen atom of a heteroarylcan be optionally oxidized to the corresponding N-oxide. Non-limitingexamples of suitable heteroaryls include pyridyl, pyrazinyl, furanyl,thienyl, pyrimidinyl, isoxazolyl, isothiazolyl, oxazolyl, thiazolyl,pyrazolyl, furazanyl, pyrrolyl, pyrazolyl, triazolyl,1,2,4-thiadiazolyl, pyrazinyl, pyridazinyl, quinoxalinyl, phthalazinyl,imidazo[1,2-a]pyridinyl, imidazo[2,1-b]thiazolyl, benzofurazanyl,indolyl, azaindolyl, benzimidazolyl, benzothienyl, quinolinyl,imidazolyl, thienopyridyl, quinazolinyl, thienopyrimidyl,pyrrolopyridyl, imidazopyridyl, isoquinolinyl, benzoazaindolyl,1,2,4-triazinyl, benzothiazolyl and the like.

“Aralkyl” or “arylalkyl” means an aryl-alkyl- group in which the aryland alkyl are as previously described. Preferred aralkyls comprise alower alkyl group. Non-limiting examples of suitable aralkyl groupsinclude benzyl, 2-phenethyl and naphthalenylmethyl. The bond to theparent moiety is through the alkyl.

“Alkylaryl” means an alkyl-aryl- group in which the alkyl and aryl areas previously described. Preferred alkylaryls comprise a lower alkylgroup. Non-limiting example of a suitable alkylaryl group is tolyl. Thebond to the parent moiety is through the aryl.

“Cycloalkyl” means a non-aromatic mono- or multicyclic ring systemcomprising about 3 to about 10 carbon atoms, preferably about 5 to about10 carbon atoms. Preferred cycloalkyl rings contain about 5 to about 7ring atoms. The cycloalkyl can be optionally substituted with one ormore “ring system substituents” which may be the same or different, andare as defined above. Non-limiting examples of suitable monocycliccycloalkyls include cyclopropyl, cyclopentyl, cyclohexyl, cycloheptyland the like. Non-limiting examples of suitable multicyclic cycloalkylsinclude 1-decalinyl, norbornyl, adamantyl and the like.

“Halogen” means fluorine, chlorine, bromine, or iodine. Preferred arefluorine, chlorine or bromine, and more preferred are fluorine andchlorine.

“Ring system substituent” means a substituent attached to an aromatic ornon-aromatic ring system which, for example, replaces an availablehydrogen on the ring system. Ring system substituents may be the same ordifferent, each being independently selected from the group consistingof aryl, heteroaryl, aralkyl, alkylaryl, heteroaralkyl, alkylheteroaryl,hydroxy, hydroxyalkyl, alkoxy, aryloxy, aralkoxy, acyl, aroyl, halo,nitro, cyano, carboxy, alkoxycarbonyl, aryloxycarbonyl,aralkoxycarbonyl, alkylsulfonyl, arylsulfonyl, heteroarylsulfonyl,alkylthio, arylthio, heteroarylthio, aralkylthio, heteroaralkylthio,cycloalkyl, heterocyclyl, Y₁Y₂N—, Y₁Y₂N-alkyl-, Y₁Y₂NC(O)— andY₁Y₂NSO₂—, wherein Y₁ and Y₂ may be the same or different and areindependently selected from the group consisting of hydrogen, alkyl,aryl, and aralkyl.

“Heterocyclyl” means a non-aromatic saturated monocyclic or multicyclicring system comprising about 3 to about 10 ring atoms, preferably about5 to about 10 ring atoms, in which one or more of the atoms in the ringsystem is an element other than carbon, for example nitrogen, oxygen orsulfur, alone or in combination. There are no adjacent oxygen and/orsulfur atoms present in the ring system. Preferred heterocyclyls containabout 5 to about 6 ring atoms. The prefix aza, oxa or thia before theheterocyclyl root name means that at least a nitrogen, oxygen or sulfuratom respectively is present as a ring atom. Any —NH in a heterocyclylring may exist protected such as, for example, as an —N(Boc), —N(CBz),—N(Tos) group and the like; such protected moieties are also consideredpart of this invention. The heterocyclyl can be optionally substitutedby one or more “ring system substituents” which may be the same ordifferent, and are as defined herein. The nitrogen or sulfur atom of theheterocyclyl can be optionally oxidized to the corresponding N-oxide,S-oxide or S,S-dioxide. Non-limiting examples of suitable monocyclicheterocyclyl rings include piperidyl, pyrrolidinyl, piperazinyl,morpholinyl, thiomorpholinyl, thiazolidinyl, 1,4-dioxanyl,tetrahydrofuranyl, tetrahydrothiophenyl, and the like.

It should be noted that in hetero-atom containing ring systems of thisinvention, there are no hydroxyl groups on carbon atoms adjacent to a N,O or S, as well as there are no N or S groups on carbon adjacent toanother heteroatom. Thus, for example, in the ring:

there is no —OH attached directly to carbons marked 2 and 5.

“Alkynylalkyl” means an alkynyl-alkyl-group in which the alkynyl andalkyl are as previously described. Preferred alkynylalkyls contain alower alkynyl and a lower alkyl group. The bond to the parent moiety isthrough the alkyl. Non-limiting examples of suitable alkynylalkyl groupsinclude propargylmethyl.

“Heteroaralkyl” means a heteroaryl-alkyl-group in which the heteroaryland alkyl are as previously described. Preferred heteroaralkyls containa lower alkyl group. Non-limiting examples of suitable aralkyl groupsinclude pyridylmethyl, and quinolin-3-ylmethyl. The bond to the parentmoiety is through the alkyl.

“Hydroxyalkyl” means a HO-alkyl- group in which alkyl is as previouslydefined. Preferred hydroxyalkyls contain lower alkyl. Non-limitingexamples of suitable hydroxyalkyl groups include hydroxymethyl and2-hydroxyethyl.

“Acyl” means an H—C(O)—, alkyl-C(O)— or cycloalkyl-C(O)—, group in whichthe various groups are as previously described. The bond to the parentmoiety is through the carbonyl. Preferred acyls contain a lower alkyl.Non-limiting examples of suitable acyl groups include formyl, acetyl andpropanoyl.

“Aroyl” means an aryl-C(O)— group in which the aryl group is aspreviously described. The bond to the parent moiety is through thecarbonyl. Non-limiting examples of suitable groups include benzoyl and1-naphthoyl.

“Alkoxy” means an alkyl-O— group in which the alkyl group is aspreviously described. Non-limiting examples of suitable alkoxy groupsinclude methoxy, ethoxy, n-propoxy, isopropoxy and n-butoxy. The bond tothe parent moiety is through the ether oxygen.

“Aryloxy” means an aryl-O— group in which the aryl group is aspreviously described. Non-limiting examples of suitable aryloxy groupsinclude phenoxy and naphthoxy. The bond to the parent moiety is throughthe ether oxygen.

“Aralkyloxy” means an aralkyl-O— group in which the aralkyl group is aspreviously described. Non-limiting examples of suitable aralkyloxygroups include benzyloxy and 1- or 2-naphthalenemethoxy. The bond to theparent moiety is through the ether oxygen.

“Alkylthio” means an alkyl-S— group in which the alkyl group is aspreviously described. Non-limiting examples of suitable alkylthio groupsinclude methylthio and ethylthio. The bond to the parent moiety isthrough the sulfur.

“Arylthio” means an aryl-S— group in which the aryl group is aspreviously described. Non-limiting examples of suitable arylthio groupsinclude phenylthio and naphthylthio. The bond to the parent moiety isthrough the sulfur.

“Aralkylthio” means an aralkyl-S— group in which the aralkyl group is aspreviously described. Non-limiting example of a suitable aralkylthiogroup is benzylthio. The bond to the parent moiety is through thesulfur.

“Alkoxycarbonyl” means an alkyl-O—CO— group. Non-limiting examples ofsuitable alkoxycarbonyl groups include methoxycarbonyl andethoxycarbonyl. The bond to the parent moiety is through the carbonyl.

“Aryloxycarbonyl” means an aryl-O—C(O)— group. Non-limiting examples ofsuitable aryloxycarbonyl groups include phenoxycarbonyl andnaphthoxycarbonyl. The bond to the parent moiety is through thecarbonyl.

“Aralkoxycarbonyl” means an aralkyl-O—C(O)— group. Non-limiting exampleof a suitable aralkoxycarbonyl group is benzyloxycarbonyl. The bond tothe parent moiety is through the carbonyl.

“Alkylsulfonyl” means an alkyl-S(O₂)— group. Preferred groups are thosein which the alkyl group is lower alkyl. The bond to the parent moietyis through the sulfonyl.

“Arylsulfonyl” means an aryl-S(O₂)— group. The bond to the parent moietyis through the sulfonyl.

The term “substituted” means that one or more hydrogens on thedesignated atom is replaced with a selection from the indicated group,provided that the designated atom's normal valency under the existingcircumstances is not exceeded, and that the substitution results in astable compound. Combinations of substituents and/or variables arepermissible only if such combinations result in stable compounds. By“stable compound” or “stable structure” is meant a compound that issufficiently robust to survive isolation to a useful degree of purityfrom a reaction mixture, and formulation into an efficacious therapeuticagent.

The term “optionally substituted” means optional substitution with thespecified groups, radicals or moieties.

It should also be noted that any heteroatom with unsatisfied valences inthe text, schemes, examples and Tables herein is assumed to have thehydrogen atom to satisfy the valences.

When a functional group in a compound is termed “protected”, this meansthat the group is in modified form to preclude undesired side reactionsat the protected site when the compound is subjected to a reaction.Suitable protecting groups will be recognized by those with ordinaryskill in the art as well as by reference to standard textbooks such as,for example, T. W. Greene et al, Protective Groups in organic Synthesis(1991), Wiley, N.Y.

When any variable (e.g., aryl, heterocycle, R², etc.) occurs more thanone time in any constituent or in Formula III, its definition on eachoccurrence is independent of its definition at every other occurrence.

As used herein, the term “composition” is intended to encompass aproduct comprising the specified ingredients in the specified amounts,as well as any product which results, directly or indirectly, fromcombination of the specified ingredients in the specified amounts.

Prodrugs and solvates of the compounds of the invention are alsocontemplated herein. The term “prodrug”, as employed herein, denotes acompound that is a drug precursor which, upon administration to asubject, undergoes chemical conversion by metabolic or chemicalprocesses to yield a compound of Formula III or a salt and/or solvatethereof. A discussion of prodrugs is provided in T. Higuchi and V.Stella, Pro-drugs as Novel Delivery Systems (1987) 14 of the A.C.S.Symposium Series, and in Bioreversible Carriers in Drug Design, (1987)Edward B. Roche, ed., American Pharmaceutical Association and PergamonPress, both of which are incorporated herein by reference thereto.

“Solvate” means a physical association of a compound of this inventionwith one or more solvent molecules. This physical association involvesvarying degrees of ionic and covalent bonding, including hydrogenbonding. In certain instances the solvate will be capable of isolation,for example when one or more solvent molecules are incorporated in thecrystal lattice of the crystalline solid. “Solvate” encompasses bothsolution-phase and isolatable solvates. Non-limiting examples ofsuitable solvates include ethanolates, methanolates, and the like.“Hydrate” is a solvate wherein the solvent molecule is H₂O.

“Effective amount” or “therapeutically effective amount” is meant todescribe an amount of compound or a composition of the present inventioneffective in inhibiting the CDK(s) and thus producing the desiredtherapeutic, ameliorative, inhibitory or preventative effect.

The compounds of Formula III can form salts which are also within thescope of this invention. Reference to a compound of Formula III hereinis understood to include reference to salts thereof, unless otherwiseindicated. The term “salt(s)”, as employed herein, denotes acidic saltsformed with inorganic and/or organic acids, as well as basic saltsformed with inorganic and/or organic bases. In addition, when a compoundof Formula III contains both a basic moiety, such as, but not limited toa pyridine or imidazole, and an acidic moiety, such as, but not limitedto a carboxylic acid, zwitterions (“inner salts”) may be formed and areincluded within the term “salt(s)” as used herein. Pharmaceuticallyacceptable (i.e., non-toxic, physiologically acceptable) salts arepreferred, although other salts are also useful. Salts of the compoundsof the Formula III may be formed, for example, by reacting a compound ofFormula III with an amount of acid or base, such as an equivalentamount, in a medium such as one in which the salt precipitates or in anaqueous medium followed by lyophilization.

Exemplary acid addition salts include acetates, ascorbates, benzoates,benzenesulfonates, bisulfates, borates, butyrates, citrates,camphorates, camphorsulfonates, fumarates, hydrochlorides,hydrobromides, hydroiodides, lactates, maleates, methanesulfonates,naphthalenesulfonates, nitrates, oxalates, phosphates, propionates,salicylates, succinates, sulfates, tartarates, thiocyanates,toluenesulfonates (also known as tosylates,) and the like. Additionally,acids which are generally considered suitable for the formation ofpharmaceutically useful salts from basic pharmaceutical compounds arediscussed, for example, by S. Berge et al, Journal of PharmaceuticalSciences (1977) 66(1) 1–19; P. Gould, International J. of Pharmaceutics(1986) 33 201–217; Anderson et al, The Practice of Medicinal Chemistry(1996), Academic Press, New York; and in The Orange Book (Food & DrugAdministration, Washington, D.C. on their website). These disclosuresare incorporated herein by reference thereto.

Exemplary basic salts include ammonium salts, alkali metal salts such assodium, lithium, and potassium salts, alkaline earth metal salts such ascalcium and magnesium salts, salts with organic bases (for example,organic amines) such as dicyclohexylamines, t-butyl amines, and saltswith amino acids such as arginine, lysine and the like. Basicnitrogen-containing groups may be quarternized with agents such as loweralkyl halides (e.g. methyl, ethyl, and butyl chlorides, bromides andiodides), dialkyl sulfates (e.g. dimethyl, diethyl, and dibutylsulfates), long chain halides (e.g. decyl, lauryl, and stearylchlorides, bromides and iodides), aralkyl halides (e.g. benzyl andphenethyl bromides), and others.

All such acid salts and base salts are intended to be pharmaceuticallyacceptable salts within the scope of the invention and all acid and basesalts are considered equivalent to the free forms of the correspondingcompounds for purposes of the invention.

Compounds of Formula III, and salts, solvates and prodrugs thereof, mayexist in their tautomeric form (for example, as an amide or iminoether). All such tautomeric forms are contemplated herein as part of thepresent invention.

All stereoisomers (for example, geometric isomers, optical isomers andthe like) of the present compounds (including those of the salts,solvates and prodrugs of the compounds as well as the salts and solvatesof the prodrugs), such as those which may exist due to asymmetriccarbons on various substituents, including enantiomeric forms (which mayexist even in the absence of asymmetric carbons), rotameric forms,atropisomers, and diastereomeric forms, are contemplated within thescope of this invention, as are positional isomers (such as, forexample, 4-pyridyl and 3-pyridyl). Individual stereoisomers of thecompounds of the invention may, for example, be substantially free ofother isomers, or may be admixed, for example, as racemates or with allother, or other selected, stereoisomers. The chiral centers of thepresent invention can have the S or R configuration as defined by theIUPAC 1974 Recommendations. The use of the terms “salt”, “solvate”“prodrug” and the like, is intended to equally apply to the salt,solvate and prodrug of enantiomers, stereoisomers, rotamers, tautomers,positional isomers, racemates or prodrugs of the inventive compounds.

The compounds according to the invention have pharmacologicalproperties; in particular, the compounds of Formula III can beinhibitors of protein kinases such as the cyclin dependent kinases(CDKs), for example, CDC2 (CDK1), CDK2, CDK4, CDK5, CDK6, CDK7 and CDK8.The novel compounds of Formula III are expected to be useful in thetherapy of proliferative diseases such as cancer, autoimmune diseases,viral diseases, fungal diseases, neurological/neurodegenerativedisorders, arthritis, inflammation, anti-proliferative (e.g., ocularretinopathy), neuronal, alopecia and cardiovascular disease. Many ofthese diseases and disorders are listed in U.S. Pat. No. 6,413,974 citedearlier, the disclosure of which is incorporated herein.

More specifically, the compounds of Formula III can be useful in thetreatment of a variety of cancers, including (but not limited to) thefollowing: carcinoma, including that of the bladder, breast, colon,kidney, liver, lung, including small cell lung cancer, esophagus, gallbladder, ovary, pancreas, stomach, cervix, thyroid, prostate, and skin,including squamous cell carcinoma;

hematopoietic tumors of lymphoid lineage, including leukemia, acutelymphocytic leukemia, acute lymphoblastic leukemia, B-cell lymphoma,T-cell lymphoma, Hodgkins lymphoma, non-Hodgkins lymphoma, hairy celllymphoma and Burkett's lymphoma;

hematopoietic tumors of myeloid lineage, including acute and chronicmyelogenous leukemias, myelodysplastic syndrome and promyelocyticleukemia;

tumors of mesenchymal origin, including fibrosarcoma andrhabdomyosarcoma;

tumors of the central and peripheral nervous system, includingastrocytoma, neuroblastoma, glioma and schwannomas; and

other tumors, including melanoma, seminoma, teratocarcinoma,osteosarcoma, xenoderoma pigmentosum, keratoctanthoma, thyroidfollicular cancer and Kaposi's sarcoma.

Due to the key role of CDKs in the regulation of cellular proliferationin general, inhibitors could act as reversible cytostatic agents whichmay be useful in the treatment of any disease process which featuresabnormal cellular proliferation, e.g., benign prostate hyperplasia,familial adenomatosis polyposis, neuro-fibromatosis, atherosclerosis,pulmonary fibrosis, arthritis, psoriasis, glomerulonephritis, restenosisfollowing angioplasty or vascular surgery, hypertrophic scar formation,inflammatory bowel disease, transplantation rejection, endotoxic shock,and fungal infections.

Compounds of Formula III may also be useful in the treatment ofAlzheimer's disease, as suggested by the recent finding that CDK5 isinvolved in the phosphorylation of tau protein (J. Biochem, (1995) 117,741–749).

Compounds of Formula III may induce or inhibit apoptosis. The apoptoticresponse is aberrant in a variety of human diseases. Compounds ofFormula III, as modulators of apoptosis, will be useful in the treatmentof cancer (including but not limited to those types mentionedhereinabove), viral infections (including but not limited to herpevirus,poxvirus, Epstein-Barr virus, Sindbis virus and adenovirus), preventionof AIDS development in HIV-infected individuals, autoimmune diseases(including but not limited to systemic lupus, erythematosus, autoimmunemediated glomerulonephritis, rheumatoid arthritis, psoriasis,inflammatory bowel disease, and autoimmune diabetes mellitus),neurodegenerative disorders (including but not limited to Alzheimer'sdisease, AIDS-related dementia, Parkinson's disease, amyotrophic lateralsclerosis, retinitis pigmentosa, spinal muscular atrophy and cerebellardegeneration), myelodysplastic syndromes, aplastic anemia, ischemicinjury associated with myocardial infarctions, stroke and reperfusioninjury, arrhythmia, atherosclerosis, toxin-induced or alcohol relatedliver diseases, hematological diseases (including but not limited tochronic anemia and aplastic anemia), degenerative diseases of themusculoskeletal system (including but not limited to osteoporosis andarthritis) aspirin-sensitive rhinosinusitis, cystic fibrosis, multiplesclerosis, kidney diseases and cancer pain.

Compounds of Formula III, as inhibitors of the CDKs, can modulate thelevel of cellular RNA and DNA synthesis. These agents would therefore beuseful in the treatment of viral infections (including but not limitedto HIV, human papilloma virus, herpesvirus, poxvirus, Epstein-Barrvirus, Sindbis virus and adenovirus).

Compounds of Formula III may also be useful in the chemoprevention ofcancer. Chemoprevention is defined as inhibiting the development ofinvasive cancer by either blocking the initiating mutagenic event or byblocking the progression of pre-malignant cells that have alreadysuffered an insult or inhibiting tumor relapse.

Compounds of Formula III may also be useful in inhibiting tumorangiogenesis and metastasis.

Compounds of Formula III may also act as inhibitors of other proteinkinases, e.g., protein kinase C, her2, raf 1, MEK1, MAP kinase, EGFreceptor, PDGF receptor, IGF receptor, PI3 kinase, wee1 kinase, Src, AbIand thus be effective in the treatment of diseases associated with otherprotein kinases.

Another aspect of this invention is a method of treating a mammal (e.g.,human) having a disease or condition associated with the CDKs byadministering a therapeutically effective amount of at least onecompound of Formula III, or a pharmaceutically acceptable salt orsolvate of said compound to the mammal.

A preferred dosage is about 0.001 to 500 mg/kg of body weight/day of thecompound of Formula III. An especially preferred dosage is about 0.01 to25 mg/kg of body weight/day of a compound of Formula III, or apharmaceutically acceptable salt or solvate of said compound.

The compounds of this invention may also be useful in combination(administered together or sequentially) with one or more of anti-cancertreatments such as radiation therapy, and/or one or more anti-canceragents selected from the group consisting of cytostatic agents,cytotoxic agents (such as for example, but not limited to, DNAinteractive agents (such as cisplatin or doxorubicin)); taxanes (e.g.taxotere, taxol); topoisomerase II inhibitors (such as etoposide);topoisomerase I inhibitors (such as irinotecan (or CPT-11), camptostar,or topotecan); tubulin interacting agents (such as paclitaxel, docetaxelor the epothilones); hormonal agents (such as tamoxifen); thymidilatesynthase inhibitors (such as 5-fluorouracil); anti-metabolites (such asmethoxtrexate); alkylating agents (such as temozolomide (TEMODAR™ fromSchering-Plough Corporation, Kenilworth, N.J.), cyclophosphamide);Farnesyl protein transferase inhibitors (such as, SARASAR™(4-[2-[4-[(11R)-3,10-dibromo-8-chloro-6,11-dihydro-5H-benzo[5,6]cyclohepta[1,2-b]pyridin-11-yl-]-1-piperidinyl]-2-oxoehtyl]-1-piperidinecarboxamide,or SCH 66336 from Schering-Plough Corporation, Kenilworth, N.J.),tipifarnib (Zarnestra® or R115777 from Janssen Pharmaceuticals),L778,123 (a farnesyl protein transferase inhibitor from Merck & Company,Whitehouse Station, N.J.), BMS 214662 (a farnesyl protein transferaseinhibitor from Bristol-Myers Squibb Pharmaceuticals, Princeton, N.J.);signal transduction inhibitors (such as, Iressa (from Astra ZenecaPharmaceuticals, England), Tarceva (EGFR kinase inhibitors), antibodiesto EGFR (e.g., C225), GLEEVEC™ (C-abl kinase inhibitor from NovartisPharmaceuticals, East Hanover, N.J.); interferons such as, for example,intron (from Schering-Plough Corporation), Peg-Intron (fromSchering-Plough Corporation); hormonal therapy combinations; aromatasecombinations; ara-C, adriamycin, cytoxan, and gemcitabine.

Other anti-cancer (also known as anti-neoplastic) agents include but arenot limited to Uracil mustard, Chlormethine, Ifosfamide, Melphalan,Chlorambucil, Pipobroman, Triethylenemelamine,Triethylenethiophosphoramine, Busulfan, Carmustine, Lomustine,Streptozocin, Dacarbazine, Floxuridine, Cytarabine, 6-Mercaptopurine,6-Thioguanine, Fludarabine phosphate, oxaliplatin, leucovirin,oxaliplatin (ELOXATIN™ from Sanofi-Synthelabo Pharmaeuticals, France),Pentostatine, Vinblastine, Vincristine, Vindesine, Bleomycin,Dactinomycin, Daunorubicin, Doxorubicin, Epirubicin, Idarubicin,Mithramycin, Deoxycoformycin, Mitomycin-C, L-Asparaginase, Teniposide17α-Ethinylestradiol, Diethylstilbestrol, Testosterone, Prednisone,Fluoxymesterone, Dromostanolone propionate, Testolactone,Megestrolacetate, Methylprednisolone, Methyltestosterone, Prednisolone,Triamcinolone, Chlorotrianisene, Hydroxyprogesterone, Aminoglutethimide,Estramustine, Medroxyprogesteroneacetate, Leuprolide, Flutamide,Toremifene, goserelin, Cisplatin, Carboplatin, Hydroxyurea, Amsacrine,Procarbazine, Mitotane, Mitoxantrone, Levamisole, Navelbene,Anastrazole, Letrazole, Capecitabine, Reloxafine, Droloxafine, orHexamethylmelamine.

If formulated as a fixed dose, such combination products employ thecompounds of this invention within the dosage range described herein andthe other pharmaceutically active agent or treatment within its dosagerange. For example, the CDC2 inhibitor olomucine has been found to actsynergistically with known cytotoxic agents in inducing apoptosis (J.Cell Sci., (1995) 108, 2897. Compounds of Formula III may also beadministered sequentially with known anticancer or cytotoxic agents whena combination formulation is inappropriate. The invention is not limitedin the sequence of administration; compounds of Formula III may beadministered either prior to or after administration of the knownanticancer or cytotoxic agent. For example, the cytotoxic activity ofthe cyclin-dependent kinase inhibitor flavopiridol is affected by thesequence of administration with anticancer agents. Cancer Research,(1997) 57, 3375. Such techniques are within the skills of personsskilled in the art as well as attending physicians.

Accordingly, in an aspect, this invention includes combinationscomprising an amount of at least one compound of Formula III, or apharmaceutically acceptable salt or solvate thereof, and an amount ofone or more anti-cancer treatments and anti-cancer agents listed abovewherein the amounts of the compounds/ treatments result in desiredtherapeutic effect.

The pharmacological properties of the compounds of this invention may beconfirmed by a number of pharmacological assays. The exemplifiedpharmacological assays which are described later have been carried outwith the compounds according to the invention and their salts.

This invention is also directed to pharmaceutical compositions whichcomprise at least one compound of Formula III, or a pharmaceuticallyacceptable salt or solvate of said compound and at least onepharmaceutically acceptable carrier.

For preparing pharmaceutical compositions from the compounds describedby this invention, inert, pharmaceutically acceptable carriers can beeither solid or liquid. Solid form preparations include powders,tablets, dispersible granules, capsules, cachets and suppositories. Thepowders and tablets may be comprised of from about 5 to about 95 percentactive ingredient. Suitable solid carriers are known in the art, e.g.,magnesium carbonate, magnesium stearate, talc, sugar or lactose.Tablets, powders, cachets and capsules can be used as solid dosage formssuitable for oral administration. Examples of pharmaceuticallyacceptable carriers and methods of manufacture for various compositionsmay be found in A. Gennaro (ed.), Remington's Pharmaceutical Sciences,18^(th) Edition, (1990), Mack Publishing Co., Easton, Pa.

Liquid form preparations include solutions, suspensions and emulsions.As an example may be mentioned water or water-propylene glycol solutionsfor parenteral injection or addition of sweeteners and opacifiers fororal solutions, suspensions and emulsions. Liquid form preparations mayalso include solutions for intranasal administration.

Aerosol preparations suitable for inhalation may include solutions andsolids in powder form, which may be in combination with apharmaceutically acceptable carrier, such as an inert compressed gas,e.g. nitrogen.

Also included are solid form preparations that are intended to beconverted, shortly before use, to liquid form preparations for eitheroral or parenteral administration. Such liquid forms include solutions,suspensions and emulsions.

The compounds of the invention may also be deliverable transdermally.The transdermal compositions can take the form of creams, lotions,aerosols and/or emulsions and can be included in a transdermal patch ofthe matrix or reservoir type as are conventional in the art for thispurpose.

The compounds of this invention may also be delivered subcutaneously.

Preferably the compound is administered orally.

Preferably, the pharmaceutical preparation is in a unit dosage form. Insuch form, the preparation is subdivided into suitably sized unit dosescontaining appropriate quantities of the active component, e.g., aneffective amount to achieve the desired purpose.

The quantity of active compound in a unit dose of preparation may bevaried or adjusted from about 1 mg to about 100 mg, preferably fromabout 1 mg to about 50 mg, more preferably from about 1 mg to about 25mg, according to the particular application.

The actual dosage employed may be varied depending upon the requirementsof the patient and the severity of the condition being treated.Determination of the proper dosage regimen for a particular situation iswithin the skill of the art. For convenience, the total daily dosage maybe divided and administered in portions during the day as required.

The amount and frequency of administration of the compounds of theinvention and/or the pharmaceutically acceptable salts thereof will beregulated according to the judgment of the attending clinicianconsidering such factors as age, condition and size of the patient aswell as severity of the symptoms being treated. A typical recommendeddaily dosage regimen for oral administration can range from about 1mg/day to about 500 mg/day, preferably 1 mg/day to 200 mg/day, in two tofour divided doses.

Another aspect of this invention is a kit comprising a therapeuticallyeffective amount of at least one compound of Formula III, or apharmaceutically acceptable salt or solvate of said compound and apharmaceutically acceptable carrier, vehicle or diluent.

Yet another aspect of this invention is a kit comprising an amount of atleast one compound of Formula III, or a pharmaceutically acceptable saltor solvate of said compound and an amount of at least one anticancertherapy and/or anti-cancer agent listed above, wherein the amounts ofthe two or more ingredients result in desired therapeutic effect.

The invention disclosed herein is exemplified by the followingpreparations and examples which should not be construed to limit thescope of the disclosure. Alternative mechanistic pathways and analogousstructures will be apparent to those skilled in the art.

Where NMR data are presented, ¹H spectra were obtained on either aVarian VXR-200 (200 MHz, ¹H), Varian Gemini-300 (300 MHz) or XL-400 (400MHz) and are reported as ppm down field from Me₄Si with number ofprotons, multiplicities, and coupling constants in Hertz indicatedparenthetically. Where LC/MS data are presented, analyses was performedusing an Applied Biosystems API-100 mass spectrometer and ShimadzuSCL-10A LC column: Altech platinum C18, 3 micron, 33 mm×7 mm ID;gradient flow: 0 min—10% CH₃CN, 5 min—95% CH₃CN, 7 min—95% CH₃CN, 7.5min—10% CH₃CN, 9 min—stop. The retention time and observed parent ionare given.

The following solvents and reagents may be referred to by theirabbreviations in parenthesis:

-   Thin layer chromatography: TLC-   dichloromethane: CH₂Cl₂-   ethyl acetate: AcOEt or EtOAc-   methanol: MeOH-   trifluoroacetate: TFA-   triethylamine: Et₃N or TEA-   butoxycarbonyl: n-Boc or Boc-   nuclear magnetic resonance spectroscopy: NMR-   liquid chromatography mass spectrometry: LCMS-   high resolution mass spectrometry: HRMS-   milliliters: mL-   millimoles: mmol-   microliters: μl-   grams: g-   milligrams: mg    room temperature or rt (ambient): about 25° C.

EXAMPLES

In general, the compounds described in this invention can be preparedthrough the general routes described below. Treatment of the startingnitrile (Scheme 1) with potassium t-butoxide and ethyl formate givesrise to the

intermediate enol 2 which upon treatment with hydrazine gives thedesired substituted 3-aminopyrazole. Condensation of the compounds oftype 3 with the appropriately functionalized keto ester of type 5 givesrise to the pyridones 6 as shown in Scheme 3. The keto esters used inthis general route are either commercially available or can be made asillustrated in Scheme 2.

The chlorides of type 9 can be prepared by treatment of the pyridones 8with POCI₃. When R² is equal to H, substitution in this position ispossible on the compounds of type 9 by electrophilic halogenation,acylation, and various other electrophilic aromatic substitutions.

Incorporation of the N7-amino functionality can be accomplished throughdisplacement of the chloride of compounds of type 9 by reaction with theappropriate amine as shown in Scheme 3.

When R³=OEt in compounds of type 6, the dichlorides of type 12 caneasily be prepared as outlined in Scheme 4. Selective displacements ofthe 7-chloride gives rise to compounds of type 13, which can readily beconverted to products of type 14.

In compounds of type 15 as shown in Scheme 5, chlorination of thesulfonic acid to give 16, followed by direct amine displacement leads tocompounds of type 17.

Preparative Example 1

Step A:

A procedure in German patent DE 19834047 A1, p 19 was followed. To asolution of KOtBu (6.17 g, 0.055 mol) in anhydrous THF (40 mL) wasadded, dropwise, a solution of cyclopropylacetonitrile (2.0 g, 0.025mol) and ethyl formate (4.07 g, 0.055 mol) in anhydrous THF (4 mL). Aprecipitate formed immediately. Stir this mixture for 12 hr. Concentrateunder vacuum and stir the residue with Et₂O (50 mL). Decant and wash theresulting residue Et₂O (2×50 mL) and remove Et₂O from the residue undervacuum. Dissolve the residue in cold H₂O (20 mL) and adjust to pH 4—5with 12 N HCl. Extract the mixture with CH₂Cl₂ (2×50 mL). Combine theorganic layers, dry over MgSO₄ and concentrate under vacuum to give thealdehyde as a tan liquid.

Step B:

The product from Preparative Example 1, Step A (2.12 g, 0.0195 mol),NH₂NH₂·H₂O (1.95 g, 0.039 mol) and 1.8 g (0.029 mole) of glacial CH₃CO₂H(1.8 g, 0.029 mol) were dissolved in EtOH (10 mL). It was refluxed for 6hr and concentrated under vacuum. The residue was slurried in CH₂Cl₂(150 mL) and the pH adjusted to 9 with 1 N NaOH. The organic layer waswashed with brine, dried over MgSO₄ and concentrated under vacuum togive the product as a waxy orange solid.

Preparative Examples 2–3

By essentially the same procedure set forth in Preparative Example 1,only substituting the nitrile shown in Column 2 of Table 2, thecompounds in Column 3 of Table 2 were prepared:

TABLE 2 Prep. Ex. Column 2 Column 3 2

3

Preparative Example 4

The reactions were done as outlined in (K. O. Olsen, J. Org. Chem.,(1987) 52, 4531–4536.). Thus, to a stirred solution of lithiumdiisopropylamide in THF at −65 to −70° C. was added freshly distilledethyl acetate, dropwise. The resulting solution was stirred for 30 minand the acid chloride was added as a solution in THF. The reactionmixture was stirred at −65 to −70° C. for 30 min and then terminated bythe addition of 1 N HCl solution. The resulting two-phased mixture wasallowed to warm to ambient temperature. The resulting mixture wasdiluted with EtOAc (100 mL) the organic layer was collected. The aqueouslayer was extracted with EtOAc (100 mL). The organic layers werecombined, washed with brine, dried (Na₂SO₄), and concentrated in vacuoto give the crude β-keto esters, which were used in the subsequentcondensations.

Preparative Examples 5–10

By essentially the same procedure set forth in Preparative Example 4only substituting the acid chlorides shown in Column 2 of Table 3, theβ-keto esters shown in Column 3 of Table 3 were prepared:

TABLE 3 Prep. Ex. Column 2 Column 3 DATA 5

Yield = 99%LCMS: MH⁺ = 223 6

Yield = 99%LCMS: MH⁺ = 253 7

Yield = 80%LCMS: MH⁺ = 261 8

Yield = 93%MH⁺ = 199 9

Yield = 93% 10

Yield = 100%

Preparative Example 11

To a solution of the acid in THF was added Et₃N, followed by isobutylchloroformate at −20 to −30° C. After the mixture was stirred for 30 minat −20 to −30° C., triethylamine hydrochloride was filtered off underargon, and the filtrate was added to the LDA-EtOAc reaction mixture(prepared as outlined in Method A) at −65 to −70° C. After addition of 1N HCl, followed by routine workup of the reaction mixture andevaporation of the solvents, the crude β-keto esters were isolated. Thecrude material was used in the subsequent condensations.

Preparative Examples 12–13.12

By essentially the same conditions set forth in Preparative Example 11only substituting the carboxylic acid shown in Column 2 of Table 4, thecompounds shown in Column 3 of Table 4 were prepared:

TABLE 4 Prep. Ex. Column 2 Column 3 DATA 12

Yield = 99%MH⁺ = 213 13

Yield = 70%MH⁺ = 275 13.10

Yield = 99MH⁺ = 199 13.11

Yield = 99MH⁺ = 334 13.12

Yield = 99MH⁺ = 334

Preparative Example 14

A solution of 3-aminopyrazole (2.0 g, 24.07 mmol) and ethylbenzoylacetate (4.58 mL, 1.1 eq.) in AcOH (15 mL) was heated at refluxfor 3 hours. The reaction mixture was cooled to room temperature andconcentrated in vacuo. The resulting solid was diluted with EtOAc andfiltered to give a white solid (2.04 g, 40% yield).

Preparative Examples 15–32.15

By essentially the same procedure set forth in Preparative Example 14only substituting the aminopyrazole shown in Column 2 of Table 5 and theester shown in Column 3 of Table 5, the compounds shown in Column 4 ofTable 5 were prepared:

TABLE 5 Prep. Ex. Column 2 Column 3 Column 4 15

16

17

18

19

20

21

22

23

24

25

26

27

28

29

30

31

32

32.10

32.11

32.12

32.13

32.14

32.15

Preparative Example 33

Ethyl benzoylacetate (1.76 mL, 1.1 eq.) and 3-amino-4-cyanopyrazole (1.0g, 9.25 mmol) in AcOH (5.0 mL) and H₂O (10 mL) was heated at reflux 72hours. The resulting solution was cooled to room temperature,concentrated in vacuo, and diluted with EtOAc. The resulting precipitatewas filtered, washed with EtOAc, and dried in vacuo (0.47 g, 21% yield).

Preparative Example 33.10

A procedure in U.S. Pat. No. 3,907,799 was followed. Sodium (2.3 g, 2eq.) was added to EtOH (150 mL) portionwise. When the sodium wascompletely dissolved, 3-aminopyrazole (4.2 g, 0.05 mol) and diethylmalonate (8.7 g, 1.1 eq.) were added and the resulting solution heatedto reflux for 3 hours. The resulting suspension was cooled to roomtemperature and filtered. The filter cake was washed with EtOH (100 mL)and dissolved in water (250 mL). The resulting solution was cooled in anice bath and the pH adjusted to 1–2 with concentrated HCl. The resultingsuspension was filtered, washed with water (100 mL) and dried undervacuum to give a white solid (4.75 g, 63% yield).

Preparative Examples 33.11–33.12

By essentially the same procedure set forth in Preparative Example 33.10only substituting the compound shown in Column 2 of Table 5.1, thecompounds shown in Column 3 of Table 5.1 are prepared:

TABLE 5.1 Prep. Ex. Column 2 Column 3 33.11

33.12

Preparative Example 34

A solution of the compound prepared in Preparative Example 14 (1.0 g,4.73 mmol) in POCl₃ (5 mL) and pyridine (0.25 mL) was stirred at roomtemperature 3 days. The resulting slurry was diluted with Et₂O,filtered, and the solid residue washed with Et₂O. The combined Et₂Owashings were cooled to 0° C. and treated with ice. When the vigorousreaction ceased, the resulting mixture was diluted with H₂O, separated,and the aqueous layer extracted with Et₂O. The combined organics werewashed with H₂O and saturated NaCl, dried over Na₂SO₄, filtered, andconcentrated to give a pale yellow solid (0.86 g, 79% yield). LCMS:MH⁺=230.

Preparative Example 35–53.14

By essentially the same procedure set forth in Preparative Example 34,only substituting the compound shown in Column 2 of Table 6, thecompounds shown in Column 3 of Table 6 were prepared:

TABLE 6 Prep. Ex. Column 2 Column 3 DATA 35

LCMS: MH⁺ = 248 36

— 37

LCMS: MH⁺ = 298 38

LCMS: MH⁺ = 196 39

LCMS: MH⁺ = 210 40

— 41

LCMS: MH⁺ = 272 42

— 43

LCMS: MH⁺ = 255 44

— 45

Yield = 65%LCMS: MH⁺ = 260 46

Yield = 35%LCMS: MH⁺ = 290 47

Yield = 32%LCMS: MH⁺ = 298 48

Yield = 45%LCMS: MH⁺ = 236 49

Yield = 100%LCMS: MH⁺ = 250 50

Yield = 88%LCMS: MH⁺ = 314 51

Yield = 43%LCMS: MH⁺ = 223 52

Yield = 30%LCMS: MH⁺ = 295 53

Yield = 98%LCMS: MH⁺ = 244 53.10

53.11

53.12

Yield = 96MH⁺ = 371 53.13

Yield = 99MH⁺ = 371 53.14

Yield = quant.MH⁺ = 236

Preparative Example 53.15

POCl₃ (62 mL) was cooled to 5° C. under nitrogen and dimethylaniline(11.4 g, 2.8 eq.) and the compound prepared in Preparative Example 33.10(4.75 g, 0.032 mol). The reaction mixture was warmed to 60° C. andstirred overnight. The reaction mixture was cooled to 30° C. and thePOCl₃ was distilled off under reduced pressure. The residue wasdissolved in CH₂Cl₂ (300 mL) and poured onto ice. After stirring 15minutes, the pH of the mixture was adjusted to 7–8 with solid NaHCO₃.The layers were separated and the organic layer was washed with H₂O(3×200 mL), dried over MgSO₄, filtered, and concentrated. The crudeproduct was purified by flash chromatography using a 50:50 CH₂Cl₂:hexanes solution as eluent to elute the dimethyl aniline. The eluent wasthen changed to 75:25 CH₂Cl₂: hexanes to elute the desired product (4.58g, 77% yield). MS: MH⁺=188.

Preparative Examples 53.16–53.17

By essentially the same procedure set forth in Preparative Example 53.15only substituting the compound in Column 2 of Table 6.10, the compoundsshown in Column 3 of Table 6.10 are prepared:

TABLE 6.10 Prep. Ex. Column 2 Column 3 53.16

53.17

Preparative Example 54

A solution of the compound prepared in Preparative Example 34 (0.10 g,0.435 mmol) in CH₃CN (3 mL) was treated with NBS (0.085 g, 1.1 eq.). Thereaction mixture was stirred at room temperature 1 hour and concentratedunder reduced pressure. The crude product was purified by flashchromatography using an 20% EtOAc in hexanes solution as eluent (0.13 g,100% yield). LCMS: MH⁺=308.

Preparative Examples 55–67.15

By essentially the same procedure set forth in Preparative Example 54only substituting the compounds shown in Column 2 of Table 7, thecompounds shown in Column 3 of Table 7 were prepared:

TABLE 7 Prep. Ex. Column 2 Column 3 CMPD 55

LCMS: MH⁺ = 326 56

LCMS: MH⁺ = 342 57

LCMS: MH⁺ = 376 58

LCMS: MH⁺ = 274 59

LCMS: MH⁺ = 288 60

LCMS: MH⁺ = 342 61

Yield = 75%LCMS: MH⁺ = 338 62

Yield = 52%LCMS: MH⁺ = 368 63

Yield = 87%LCMS: MH⁺ = 376 64

Yield = 100%LCMS: MH⁺ = 316 65

Yield = 92%LCMS: MH⁺ = 330 66

Yield = 82%LCMS: MH⁺ = 395 67

Yield = 100%LCMS: MH⁺ = 322 67.10

67.11

67.12

Yield = 99%MH⁺ = 449 67.13

Yield = 95%MH⁺ = 449 67.14

MH⁺ = 266 67.15

Yield = quant.MH⁺ = 314

Preparative Example 68

A solution of the compound prepared in Preparative Example 35 (0.3 g,1.2 mmol) in CH₃CN (15 mL) was treated with NCS (0.18 g, 1.1 eq.) andthe resulting solution heated to reflux 4 hours. Additional NCS (0.032g, 0.2 eq.) added and the resulting solution was stirred at refluxovernight. The reaction mixture was cooled to room temperature,concentrated in vacuo and the residue purified by flash chromatographyusing a 20% EtOAc in hexanes solution as eluent (0.28 g, 83% yield).LCMS: MH⁺=282.

Preparative Example 69

By essentially the same procedure set forth in Preparative Example 68only substituting the compound shown in Column 2 of Table 8, thecompound shown in Column 3 of Table 7 was prepared:

TABLE 8 Prep. Ex. Column 2 Column 3 DATA 69

Yield = 82%LCMS: MH⁺ = 286

Example 1

The product from Preparative Example 56 (0.12 g, 0.35 mmol) and4-methylsulfonylaniline hydrochloride (0.065 g, 0.9 eq) and iPr₂NEt (1.0mL) were heated to 100° C. for 48 hours. The reaction mixture was cooledto room temperature and purified by Preparative thin layerchromatography using a 5% (10% NH₄OH in MeOH) solution in CH₂Cl₂ aseluent (0.033 g, 23% yield). LCMS: MH⁺=477. mp=180–182° C.

Examples 2–21.15

By essentially the same procedure set forth in Example 1 onlysubstituting the compound shown in Column 2 of Table 9 and the amineshown in Column 3 of Table 9, the compounds shown in Column 4 of Table 9were prepared and are prepared:

TABLE 9 Ex. Column 2 Column 3 Column 4 DATA 2

LCMS:MH⁺ =346;mp =58–65° C. 3

LCMS:MH⁺ =339;mp =112–116° C. 4

LCMS:MH⁺ =417;mp =232–235° C. 5

LCMS:MH⁺ =461;mp =117–118° C. 6

LCMS:MH⁺ =511;mp =210–212° C. 7

LCMS:MH⁺ =409;mp =214–215° C. 8

LCMS:MH⁺ =331;mp =166–168° C. 9

LCMS:MH⁺ =345;mp =144° C. 10

LCMS:MH⁺ =405;mp =210–211° C. 11

LCMS:MH⁺ =407;mp =213–216° C. 12

LCMS:MH⁺ =477;mp =249–253° C. 13

Yield =72%LCMS:MH⁺ =518. 14

Yield =75%LCMS:MH⁺ =429. 15

Yield =99%LCMS:MH⁺ =395 16

Yield =45%LCMS:MH⁺ =497. 17

18

19

20

LCMS:MH⁺ =393;mp =192–194° C. 21

LCMS:M2H⁺ =445;mp =202–204° C. 21.10

21.11

21.12

21.13

21.14

MH⁺ =401 21.15

Additional data for select examples is shown below:

Example 13

¹H NMR (CDCl₃) δ 8.21 (s, 1H), 8.07 (s, 1H), 7.66–7.64 (m, 2H),7.60–7.39 (m, 3H), 7.10–7.07 (m, 2H), 6.56 (s, 1H), 3.99 (dd, J=5.1, 4.5Hz, 4H), 3.31 (dd, J=5.1, 4.5 Hz, 4H).

Example 14

¹H NMR (CDCl₃) δ 8.16(s, 1H),8.14(d, J=2.1 Hz, 1H), 7.63 (m, 1H),7.5–7.45 (m,2H), 7.23–7.09(m,3H), 6.84–6.76 (m, 2H),6.64 (m, 1 H), 4.03(s, 3H).

Example 15

¹H NMR (CDCl₃)δ8.32(s,1H),7.51(d,1H), 7.43–7.33(m, 4H), 6.78(d,2H),6.72(t, 1H), 2.52(s,3H).

Example 16

¹H NMR (CD₃OD) δ 8.31 (s, 1H), 7.75–7.69 (m, 2H),7.64–7.60 (m,2H),7.56–7.37 (m, 2H),6.37 (s, 1H), 4.79 (s, 2H).

Example 22

Anhydrous DMF (80 mL) was added under N₂ to a mixture of sulfanilic acid(3.10 g, 17.9 mmol) and NaH (60% in mineral oil, 1.43 g, 35.8 mmol), themixture was stirred at 25° C. for 2 hr, then the product fromPreparative Example 54 (5.00 g, 16.2 mmol) was added. The mixture wasstirred at 25° C. for 24 hr, the solvent was then evaporated and theresidue was purified by chromatography on silica gel using EtOAc:MeOH(4:1) as eluent to yield pale yellow solid (2.32 g, 32% yield). LCMS:MH⁺=447. mp>250° C.

Examples 23–26

By essentially the same procedure set forth in Example 22 onlysubstituting the compound shown in Column 2 of Table 10 and the amineshown in column 3 of Table 10, the compounds shown in Column 4 of Table10 were prepared.

TABLE 10 Ex. Column 2 Column 3 Column 4 DATA 24

LCMS:M2H⁺ =445;mp =206–208° C. 25

LCMS:M2H⁺ =465;mp >250° C. 26

LCMS:MH⁺ =395;mp >250° C.

Example 27

The product from Example 22 (44 mg, 0.10 mmol), and PCl₅ (21 mg, 0.10mmol) in anhydrous 1,2-dichloroethane were stirred and refluxed under N₂for 2.5 hr. The mixture was cooled to 25° C., propylamine (0.20 mL, 2.4mmol) was added, and the mixture was stirred at 25° C. for 2 hr. Thesolvent was then evaporated and the residue was purified bychromatography on silica gel using CH₂Cl₂:EtOAc (20:1) as eluent toyield pale yellow solid (26 mg, 54% yield). LCMS: MH⁺=486. mp=201–203°C.

Examples 28–67

By essentially the same procedure set forth in Example 27 onlysubstituting the compound shown in Column 2 of Table 11 and the amineshown in Column 3 of Table 11, the compounds shown in Column 4 of Table11 were prepared:

TABLE 11 Ex. Column 2 Column 3 Column 4 DATA 28

LCMS: M2H⁺ =474; mp = 101–104° C. 29

LCMS: M2H⁺ =472; mp = 237–239° C. 30

LCMS: M⁺ =488; mp = 175–177° C. 31

NH₄OH

LCMS: MH⁺ =444; mp = 206–208° C. 32

LCMS: MH⁺ =458; mp = 231–233° C. 33

LCMS: MH⁺ =589; mp = 195–197° C. 34

LCMS: MH⁺ =717; 35

LCMS: MH⁺ =536; mp = 216–218° C. 36

LCMS: M⁺ =502; mp = 165–168° C. 37

LCMS: MH⁺ =522; mp = 147–150° C. 38

LCMS: M2H⁺ =523; mp = 192–195° C. 39

LCMS: MH⁺ =629; mp = 127–129° C. 40

LCMS: MH⁺ =514; mp >200° C. (dec.) 41

LCMS: M2H⁺ =504; mp >200° C. (dec.) 42

LCMS: M2H⁺ =504; mp = 172–173° C. 43

LCMS: MH⁺ =517; mp = 165–167° C. 44

LCMS: M2H⁺ =537; mp = 101–103° C. 45

LCMS: M2H⁺ =537; mp = 110–114° C. 46

LCMS: M2H⁺ =559; mp = 47

LCMS: M⁺ =532; mp = 90–92° C. 48

LCMS: MH⁺ =559; mp = 163–165° C. 49

LCMS: M⁺ =552; mp = 206–208° C. 50

LCMS: MH⁺ =520; mp = 122–124° C. 51

LCMS: M⁺ =532; mp = 98–100° C. 52

LCMS: M2H⁺ =518; mp = 182–184° C. 53

LCMS: MH⁺ =531; mp = 78–80° C. 54

LCMS: MH⁺ =529; mp = 228–230° C. 55

LCMS: MH⁺ =580; mp = 108–110° C. 56

LCMS: MH⁺ =580; mp = 102–105° C. 57

LCMS: M2H⁺ =528; mp = 58

LCMS:M2H+ = 557m.p. = 204–207 59

LCMS:M2H+ = 557 60

LCMS:M2H+ = 571m.p. = 114–117 61

LCMS:M2H+ = 554m.p. = 127–130 62

LCMS:M2H+ = 613m.p. = 145–149 63

LCMS:M2H+ = 613m.p. = 137–140

64

LCMS: M2H⁺ =490; mp = 87–90° C. 65

LCMS: M2H⁺ =504; mp = 115–120° C. 66

LCMS: M2H⁺ =508; mp = 67

LCMS: MH⁺ =465; mp = 99–101° C.

Example 68

Trifluoroacetic acid (2.0 mL) was added at 0° C. to a solution of theproduct from Preparative Example 33 (200 mg, 0.34 mmol) in anhydrousCH₂Cl₂. The mixture was stirred at 0° C. for 5 min, then at 25° C. for90 min, and then it was poured onto solid Na₂CO₃ (10.0 g). H₂O (150 mL)was added and the mixture was extracted with CH₂Cl₂ (3×25 mL). Theextracts were dried over Na₂SO₄, filtered, and the solvent wasevaporated. The residue was purified by chromatography on silica gelusing CH₂Cl₂:MeOH:conc. NH₄OH (10:1:0.1) as eluent to yield pale yellowsolid (100 mg, 60% yield). LCMS: M⁺=487. mp=110–112° C.

Example 69

To a solution of the compound prepared in Example 21.14 (0.10 g, 0.25mmol) and prolinol (0.12 mL, 5 eq.) and iPr₂NEt (0.22 mL, 5 eq.) washeated to reflux 24 hours. (Yield: 0.09 g, 80%). MS: MH⁺=466; m.p.=177–180° C.

Example 70–78

By essentially the same procedure set forth in Example 69 onlysubstituting the amine shown in Column 2 of Table 12, the compoundsshown in Column 3 of Table 12 are prepared:

TABLE 12 Ex. Column 2 Column 3 70

71

72

73

74

75

76

77

78

Example 79

To a solution of the compound prepared in Example 21.12 in anhydrousacetonitrile is added TMSI (4 eq.), dropwise at ambient temperature.After 10 minutes the acetonitrile is removed in vacuo. The resultingyellow foam is treated with 2 N HCl solution (7 mL) and then washedimmediately with Et₂O (5×). The pH of the aqueous is adjusted to 10 with50% NaOH (aq) and the product is isolated by saturation of the solutionwith NaCl (s) followed by extraction with CH₂Cl₂ (5×) to give thedesired product.

Example 80

By essentially the same procedure set forth in Example 79 onlysubstituting the compounds shown in Column 2 of Table 13, the compoundsshown in Column 3 of Table 13 are prepared.

TABLE 13 Ex. Column 2 Column 3 80

ASSAY

BACULOVIRUS CONSTRUCTIONS: Cyclin E was cloned into pVL1393 (Pharmingen,La Jolla, Calif.) by PCR, with the addition of 5 histidine residues atthe amino-terminal end to allow purification on nickel resin. Theexpressed protein was approximately 45 kDa. CDK2 was cloned into pVL1393by PCR, with the addition of a haemaglutinin epitope tag at thecarboxy-terminal end (YDVPDYAS). The expressed protein was approximately34 kDa in size.

ENZYME PRODUCTION: Recombinant baculoviruses expressing cyclin E andCDK2 were co-infected into SF9 cells at an equal multiplicity ofinfection (MOI=5), for 48 hrs. Cells were harvested by centrifugation at1000 RPM for 10 minutes, then pellets lysed on ice for 30 minutes infive times the pellet volume of lysis buffer containing 50 mM Tris pH8.0, 150 mM NaCl, 1% NP40, 1 mM DTT and protease inhibitors (RocheDiagnostics GmbH, Mannheim, Germany). Lysates were spun down at 15000RPM for 10 minutes and the supernatant retained. 5 ml of nickel beads(for one liter of SF9 cells) were washed three times in lysis buffer(Qiagen GmbH, Germany). Imidazole was added to the baculovirussupernatant to a final concentration of 20 mM, then incubated with thenickel beads for 45 minutes at 4° C. Proteins were eluted with lysisbuffer containing 250 mM imidazole. Eluate was dialyzed overnight in 2liters of kinase buffer containing 50 mM Tris pH 8.0, 10 mM DTT, 10 mMMgCl2, 100 uM sodium orthovanadate and 20% glycerol. Enzyme was storedin aliquots at −70° C.

IN VITRO KINASE ASSAY: Cyclin E/CDK2 kinase assays were performed in lowprotein binding 96-well plates (Corning Inc, Corning, N.Y.). Enzyme wasdiluted to a final concentration of 50 μg/ml in kinase buffer containing50 mM Tris pH 8.0, 10 mM MgCl₂, 1 mM DTT, and 0.1 mM sodiumorthovanadate. The substrate used in these reactions was a biotinylatedpeptide derived from Histone H1 (from Amersham, UK). The substrate wasthawed on ice and diluted to 2 μM in kinase buffer. Compounds werediluted in 10% DMSO to desirable concentrations. For each kinasereaction, 20 μl of the 50 μg/ml enzyme solution (1 μg of enzyme) and 20μl of the 2 μM substrate solution were mixed, then combined with 10 μlof diluted compound in each well for testing. The kinase reaction wasstarted by addition of 50 μl of 2 μM ATP and 0.1 μCi of 33P-ATP (fromAmersham, UK). The reaction was allowed to run for 1 hour at roomtemperature. The reaction was stopped by adding 200 μt of stop buffercontaining 0.1% Triton X-100, 1 mM ATP, 5 mM EDTA, and 5 mg/mlstreptavidine coated SPA beads (from Amersham, UK) for 15 minutes. TheSPA beads were then captured onto a 96-well GF/B filter plate(Packard/Perkin Elmer Life Sciences) using a Filtermate universalharvester (Packard/Perkin Elmer Life Sciences.). Non-specific signalswere eliminated by washing the beads twice with 2M NaCl then twice with2 M NaCl with 1% phosphoric acid. The radioactive signal was thenmeasured using a TopCount 96 well liquid scintillation counter (fromPackard/Perkin Elmer Life Sciences).

IC₅₀ DETERMINATION: Dose-response curves were be plotted from inhibitiondata generated, each in duplicate, from 8 point serial dilutions ofinhibitory compounds. Concentration of compound was plotted against %kinase activity, calculated by CPM of treated samples divided by CPM ofuntreated samples. To generate IC₅₀ values, the dose-response curveswere then fitted to a standard sigmoidal curve and IC₅₀ values werederived by nonlinear regression analysis. The thus-obtained IC₅₀ valuesfor some of the compounds of the invention are shown in Table 14.

TABLE 14 Ex- am- IC₅₀ Compound ple (μM)

2 0.51

1 0.41.4

3 0.042

22 0.082

28 0.080

30 0.029

31 0.045

32 0.057

38 0.040

42 0.070

43 0.034

47 0.034

48 0.025

50 0.030

53 0.011

As demonstrated above by the assay values, the compounds of the presentinvention exhibit excellent CDK inhibitory properties.

While the present invention has been described with in conjunction withthe specific embodiments set forth above, many alternatives,modifications and other variations thereof will be apparent to those ofordinary skill in the art. All such alternatives, modifications andvariations are intended to fall within the spirit and scope of thepresent invention.

1. A compound represented by the structural formula:

or a pharmaceutically acceptable salt of said compound, wherein: R is anunsubstituted aryl or aryl substituted with one or more moieties whichmoieties can be the same or different, each moiety being independentlyselected from the group consisting of halogen, CN, —OR⁵, SR⁵, —CH₂OR⁵,—C(O)R⁵, —SO₃H, —S(O₂)R⁶, —S(O₂)NR⁵R⁶, —NR⁵R⁶, —C(O)NR⁵R⁶, —CF₃, —OCF₃and heterocyclyl; R² is selected from the group consisting of R⁹, alkyl,alkynyl, alkynylalkyl, cycloalkyl, —CF₃, —C(O₂)R⁶, aryl, arylalkyl,heteroarylalkyl, heterocyclyl, alkyl substituted with 1–6 R⁹ groupswhich groups can be the same or different with each R⁹ beingindependently selected, aryl substituted with 1–3 aryl or heteroarylgroups which can be the same or different and are independently selectedfrom phenyl, pyridyl, thiophenyl, furanyl and thiazolo groups,

R³ is selected from the group consisting of H, halogen, —NR⁵R⁶,—C(O)NR⁵R⁶, alkyl, alkynyl, cycloalkyl, aryl, arylalkyl, heterocyclyl,heteroaryl and heteroarylalkyl,

wherein each of said alkyl, cycloalkyl, aryl, arylalkyl, heterocyclyl,heteroaryl and heteroarylalkyl for R³ and the heterocyclyl moietieswhose structures are shown immediately above for R³ can be substitutedor optionally independently substituted with one or more moieties whichcan be the same or different, each moiety being independently selectedfrom the group consisting of halogen, alkyl, aryl, cycloalkyl, CF₃, CN,—OCF₃, —(CR⁴R⁵)_(n)OR⁵, —OR⁵, —NR⁵R⁶, —(CR⁴R⁵)_(n)NR⁵R⁶, —C(O₂)R⁵,—C(O)R⁵, —C(O)NR⁵R⁶, —SR⁶, —S(O₂)R⁶, —S (O₂)NR⁵R⁶, —N(R⁵)S(O₂)R⁷,—N(R⁵)C(O)R⁷ and —N(R⁵)C(O)NR⁵R⁶; R⁴ is H, halo or alkyl; R⁵ is H oralkyl; R⁶ is selected from the group consisting of H, alkyl, aryl,arylalkyl, cycloalkyl, heterocyclyl, heteroaryl, and heteroarylalkyl,wherein each of said alkyl, aryl, arylalkyl, cycloalkyl, heterocyclyl,heteroaryl, and heteroarylalkyl can be unsubstituted or optionallysubstituted with one or more moieties which can be the same ordifferent, each moiety being independently selected from the groupconsisting of halogen, alkyl, aryl, cycloalkyl, CF₃, OCF₃, CN, —OR⁵,—NR⁵R¹⁰, —N(R⁵)Boc, —(CR⁴R⁵)_(n)OR⁵, —C(O₂)R⁵, —C(O)R⁵, —C(O)NR⁵R¹⁰,—SO₃H, —SR¹⁰, —S(O₂)R⁷, —S(O₂)NR⁵R¹⁰, —N(R⁵)S(O₂)R⁷, —N(R⁵)C(O)R⁷ and—N(R⁵)C(O)NR⁵R¹⁰; R¹⁰ is selected from the group consisting of H, alkyl,aryl, arylalkyl, cycloalkyl, heterocyclyl, heteroaryl, andheteroarylalkyl, wherein each of said alkyl, aryl, arylalkyl,cycloalkyl, heterocyclyl, heteroaryl, and heteroarylalkyl can beunsubstituted or optionally substituted with one or more moieties whichcan be the same or different, each moiety being independently selectedfrom the group consisting of halogen, alkyl, aryl, cycloalkyl, CF₃,OCF₃, CN, —OR⁵, —NR⁴R⁵, —N(R⁵)Boc, —(CR⁴R⁵)_(n)OR⁵, —C(O₂)R⁵,—C(O)NR⁴R⁵, —C(O)R⁵, —SO₃H, —SR⁵, —S(O₂)R⁷, —S(O₂)NR⁴R⁵, —N(R⁵)S(O₂)R⁷,—N(R⁵)C(O)R⁷ and —N(R⁵)C(O)NR⁴R⁵; or optionally (i) R⁵ and R¹⁰ in themoiety —NR⁵R¹⁰, or (ii) R⁵ and R⁶ in the moiety —NR⁵R⁶, may be joinedtogether to form a cycloalkyl or heterocyclyl moiety, with each of saidcycloalkyl or heterocyclyl moiety being unsubstituted or optionallyindependently being substituted with one or more R⁹ groups; R⁷ isselected from the group consisting of alkyl, cycloalkyl, aryl,heteroaryl, arylalkyl and heteroarylalkyl, wherein each of said alkyl,cycloalkyl, heteroarylalkyl, aryl, heteroaryl and arylalkyl can beunsubstituted or optionally independently substituted with one or moremoieties which can be the same or different, each moiety beingindependently selected from the group consisting of halogen, alkyl,aryl, cycloalkyl, CF₃, OCF₃, CN, —OR⁵, —NR⁵R¹⁰, —CH₂OR⁵, —C(O₂)R⁵,—C(O)NR⁵R¹⁰, —C(O)R⁵, —SR¹⁰, —S(O₂)R¹⁰, —S(O₂)NR⁵R¹⁰, —N(R⁵)S(O₂)R¹⁰,—N(R⁵)C(O)R¹⁰ and —N(R⁵)C(O)NR⁵ ¹⁰; R⁸ is selected from the groupconsisting of R⁶, —C(O)NR⁵R¹⁰, —S(O₂)NR⁵R¹⁰, —C(O)R⁷and —S(O₂)R⁷; R⁹ isselected from the group consisting of halogen, CN, —NR⁵R¹⁰, —C(O₂)R⁶,—C(O)NR⁵R¹⁰, —OR⁶, —SR⁶, —S(O₂)R⁷, —S(O₂)NR⁵R¹⁰, —N(R⁵)S(O₂)R⁷,—N(R⁵)C(O)R⁷ and —N(R⁵)C(O)NR⁵R¹⁰; m is 0 to 4, and n is 1 to 4, withthe following provisos: (i) that when R is an unsubstituted phenyl, thenR² is not alkyl, —C(O₂)R⁶, aryl or cycloalkyl, and (ii) that when R is aphenyl substituted with a hydroxyl group, then R² is halogen only. 2.The compound of claim 1, wherein R is an unsubstituted aryl or arylsubstituted with one or moieties which moieties can be the same ordifferent with each moiety being independently selected from the groupconsisting of halogen, CN, —OR⁵, —S(O₂)NR⁵R⁶, —SO₃H, CH₂OR⁵, —S(O₂)R⁶,—C(O)NR⁵R⁶, —CF₃, —OCF₃ and heterocyclyl; R² is halogen, CF₃, CN, loweralkyl and cycloalkyl; R³ is H, unsubstituted aryl, unsubstitutedheteroaryl, aryl substituted with one or more moieties selected from thegroup consisting of halogen, CN, —OR⁵, CF₃, —OCF₃, lower alkyl andcycloalkyl, heterocyclyl, heteroaryl substituted with one or moremoieties selected from the group consisting of halogen, CN, —OR⁵, CF₃,—OCF₃, alkyl and cycloalkyl,

R⁴ is H or lower alkyl; R⁵ is H or lower alkyl; m is 0 to 2; and n is 1or
 2. 3. The compound of claim 2, wherein R is an unsubstituted phenyl.4. The compound of claim 2, wherein R is phenyl substituted with one ormore moieties selected from the group consisting of F, Cl, Br, CN,—SO₃H, —S(O₂)NR⁵R⁶, —S(O₂)CH₃, —NR⁵R¹⁰, —OH, hydroxymethyl, CF₃ andmorpholinyl.
 5. The compound of claim 2, wherein R² is F, Cl, Br, CF₃,lower alkyl and cycloalkyl.
 6. The compound of claim 2, wherein R³ is H,lower alkyl, unsubstituted aryl, aryl substituted with one or moremoieties which can be the same or different, each moiety beingindependently selected from the group consisting of F, Cl, Br, CF₃,lower alkyl, methoxy, and CN, or


7. The compound of claim 6, wherein said lower alkyl is methyl, ethyl,isopropyl or tert-butyl.
 8. The compound of claim 2, wherein R⁴ is H. 9.The compound of claim 2, wherein R⁵ is H.
 10. The compound of claim 2,wherein m is
 0. 11. The compound of claim 2, wherein R is4-(methylsulfonyl)phenyl.
 12. The compound of claim 2, wherein R² is Cl,Br, isopropyl, ethyl, cyclopropyl, cyclobutyl or cyclopentyl.
 13. Thecompound of claim 6, wherein R³ is unsubstituted phenyl, or phenylsubstituted with —S(O₂)NR⁵R⁶.
 14. The compound of claim 6, wherein R³ istert-butyl or isopropyl.
 15. The compound of claim 6, wherein R³ is2-fluorophenyl, 2-chlorophenyl, 2,3-dichlorophenyl, 2-methylphenyl,2-methoxyphenyl,


16. The compound of claim 6, wherein R³ is 3-(trifluoromethyl)phenyl.17. The compound of claim 15, wherein R³ is:


18. The compound of claim 17, wherein R⁸ is (CH₂)_(n)OH or(CH₂)_(n)OCH₃, where n is 1 or
 2. 19. A compound selected from the groupconsisting of:

or a pharmaceutically acceptable salt thereof.
 20. A compound of theformula:

or a pharmaceutically acceptable salt thereof.
 21. A compound of claim1, in isolated and purified form.